Prediction and analyzes of non coding RNA sequences in the pea aphid genome

Small non-coding RNAs (sncRNAs) have been abundantly described as strongly implicated in the post-transcriptional regulation of transcribed units in eukaryote genomes. Silencing of genes, pseudogenes or transposable elements can be operated in the germline or in the soma through microRNA (miRNAs), PIWI associated RNAs (piRNAs) or endogenous short interfering RNAs (esiRNAs or siRNAs). MiRNAs play crucial roles in regulation of gene expression in eukaryotes. In insects, miRNAs have been shown to be involved in multiple mechanisms such as embryonic development, tissue differentiation, metamorphosis or circadian rhythm. PiRNAs (25-30 nucleotides) have been demonstrated as involved in the silencing of transposable elements (TEs) through an amplification loop in the germline while endo-siRNAs (21 nucleotides) were also involved in the silencing of TEs but in the somatic cells. All these sncRNAs are thus sculpting the whole functional transcriptome of an organism. Aphids are herbivorous insects characterised by the unique ability to feed exclusively on phloem sap. The serious damage caused by aphids is partly due to their amazing ability to adapt to environmental variations. This adaptive capacity is largely explained by their phenotypic plasticity that allows the production of distinct phenotypic morphs in response to environmental changes. This is illustrated by the switch of reproductive mode in response to seasonal changes: parthenogenetic females are produced in spring and summer whereas sexual females and males are produced in autumn. Since several years, our lab deciphers the molecular bases of genome functioning during phenotypic plasticity of the reproductive mode. These researches are based on the pea aphid Acyrthosiphon pisum for which the International Aphid Genomics Consortium developed a genome sequence and other genomic resources. The understanding of phenotypic plasticity is searched by characterizing the different steps of gene regulation, from epigenetic regulation to protein synthesis. As sncRNAs are regulators of these different steps, we decided to characterize sncRNAs in the pea aphid and to identify there putative roles in regulating phenotypic plasticity of the reproductive mode. First, we undertook deep sequencing of sncRNAs from a mixed generation sample of parthenogenetic female pea aphids. Approximately 3 million sequences were generated, corresponding to approximately 850,000 unique sequences. Then, we set protocols to use these sequences comparatively to the genome and annotations (genes and TEs) of the pea aphid Acyrthosiphon pisum, in order to create a first catalogue of miRNA and piRNA. In this presentation, we will emphasize on the methods we used for annotating these sncRNAs, and we will give some examples of some miRNAs regulated during the switch of reproductive mode. Finally, we will describe a protocol in progress to identify putative miRNA targets by combining bioinformatics approach and expression data.